Sequencing and Bioinformatics Service, Foundation for the Promotion of Health and Biomedical Research of Valencia Region, FISABIO, Valencia, Spain.
CIBER in Epidemiology and Public Health (CIBERESP), Instituto de Salud Carlos III, Madrid, Spain.
BMC Genomics. 2023 Feb 10;24(1):69. doi: 10.1186/s12864-023-09169-9.
Bacillus Calmette-Guérin (BCG) remains the only vaccine to prevent tuberculosis (TB) during childhood, with relatively low to no efficacy against pulmonary TB in adolescents and adults. BCG consists of close to 15 different substrains, where genetic variations among them might contribute to the variable protective efficacy afforded against pulmonary TB. We have shown that the vaccine candidate, BCGΔBCG1419c, which is based on BCG Pasteur, improved protection against chronic TB in murine models, as well as against pulmonary and extrapulmonary TB in guinea pigs. Here, to confirm deletion of the BCG1419c gene and to detect possible genetic variations occurring as a consequence of the spontaneous mutations that may arise during in vitro culture of mycobacteria, the genomes of BCG Pasteur ATCC 35734 and its isogenic derivative, BCGΔBCG1419c, were sequenced and subjected to a comparative analysis between them and against BCG Pasteur 1173P2.
The complete catalog of variants in genes relative to the reference genome BCG Pasteur 1173P2 (GenBank NC008769) showed that the parental strain BCG Pasteur ATCC 35734, from which the mutant BCGΔBCG1419c originated, showed five synonymous mutations, three missense mutations, and five codon insertions, whereas the BCGΔBCG1419c mutant reported the same changes. When BCG Pasteur ATCC 35734 and BCGΔBCG1419c were compared, we confirmed that the latter was devoid of the BCG1419c gene, with only one unanticipated SNP at position 2, 828, 791 which we consider has no role in vaccine properties reported thus far.
We provide evidence that the mutagenesis performed to remove BCG1419c from BCG Pasteur ATCC 35734 solely deleted this gene, and that compared with the reference strain BCG Pasteur 1173P2, few changes were present confirming that they are BCG Pasteur strains, and that changes in immunogenicity or efficacy observed thus far in BCGΔBCG1419c are most likely derived solely from the elimination of the BCG1419c gene.
卡介苗（BCG）仍然是预防儿童结核病（TB）的唯一疫苗，对青少年和成人的肺结核（TB）的疗效相对较低或没有。BCG 由近 15 种不同的亚种组成，其中它们之间的遗传变异可能导致对肺结核提供的保护效果不同。我们已经表明，基于卡介苗 Pasteur 的候选疫苗 BCGΔBCG1419c 改善了对慢性 TB 的保护作用，并且在豚鼠中对肺外和肺外 TB 也有保护作用。在这里，为了确认 BCG1419c 基因的缺失，并检测由于分枝杆菌体外培养过程中可能发生的自发突变而产生的可能的遗传变异，对卡介苗 Pasteur ATCC 35734 及其同源衍生菌 BCGΔBCG1419c 的基因组进行了测序，并对它们进行了测序。与参考基因组 BCG Pasteur 1173P2（GenBank NC008769）进行了比较分析。
相对于参考基因组 BCG Pasteur 1173P2（GenBank NC008769）的基因完整变体目录显示，亲本菌株卡介苗 Pasteur ATCC 35734，突变株 BCGΔBCG1419c 由此而来，显示出五个同义突变，三个错义突变和五个密码子插入，而突变株 BCGΔBCG1419c 则报告了相同的变化。当比较卡介苗 Pasteur ATCC 35734 和 BCGΔBCG1419c 时，我们确认后者缺乏 BCG1419c 基因，仅在位置 2,828,791 处存在一个意外的 SNP，我们认为该 SNP 在此前报道的疫苗特性中没有作用。
我们提供的证据表明，从卡介苗 Pasteur ATCC 35734 中去除 BCG1419c 所进行的诱变仅删除了该基因，并且与参考菌株 BCG Pasteur 1173P2 相比，存在的变化很少，这证实了它们是卡介苗菌株，并且迄今为止在 BCGΔBCG1419c 中观察到的免疫原性或疗效变化很可能仅源自 BCG1419c 基因的消除。
